Proc. Natl. Acad. Sci. USA doi:10.1073/pnas.1614196114 (2017)

Supplying a multi-enzyme polyketide synthase (PKS) complex with unnatural monomer units can, under certain conditions, facilitate the production of novel polyketide natural product analogs. Using several different extender units, including one containing fluorine, and a variety of multi-domain enzymatic constructs, Ad et al. have explored the mechanisms controlling the incorporation of alternate moieties into the polyketide backbone of erythromycin by the 6-deoxyerythronolide B synthase (DEBS) PKS complex. While inactivation of a cis-acyltransferase (AT) domain in DEBS allowed the incorporation of non-native extender units by removing the enzyme's selectivity for the native substrate, it also caused a switch in the mechanism of C–C bond formation that untethered the growing polyketide chain from its carrier protein, leading to premature chain termination and product release. However, complementation of the inactive DEBS cis-AT with an orthogonal trans-AT restored the canonical PKS mechanism and enabled further chain extension following the incorporation of alternative extender units. Use of this engineered hybrid PKS system enabled the efficient production of polyketides with site-specific incorporation of fluorine, such as mono- and bisfluorinated triketide lactones. This detailed understanding of the mechanisms of elongation in an engineered PKS will enable the biosynthesis of novel polyketides, though it remains to be seen how well these insights will translate to other PKS complexes.